Catechins are polyphenolic plant metabolites which belong to the flavonoid family. The molecular formula and weight of catechins are C15H14O6 and 290 g/mol. Catechin and epicatechin are epimers, with (−)-epicatechin and (+)-catechin being the most common optical isomers found in nature.
Procyanidins or condensed tannins are flavonoid oligomers whose building blocks are (+)-catechin and (−)-epicatechin. They are oligomeric end products of the flavonoid biosynthetic pathway and are now identified and recognised for their beneficial effects in human beings. They are present abundantly in the plant kingdom in fruits, barks, leaves and seeds where they provide protection against light, oxidation and predators. Procyanidins are found in many plants, mainly apples, pine bark, cinnamon bark, litchi pericarp, peanuts, grape seed, cocoa, grape skin, bilberry, cranberry, black currant, green tea and black tea.
Based on the linkage between the successive monomeric units, procyanidins are classified as Types A, B or C polyphenols.
Generally the linkage between successive monomeric units of procyanidins is between the 4th position of the ‘upper’ unit and the 8th position of the ‘lower’ unit, leading to a Type B procyanidin. Alternatively, the linkage can occur between C4 of the ‘upper’ unit and C6 of the lower unit, leading to a Type C procyanidin. Type B and C polyphenols are abundantly seen in many botanical sources. When successive monomeric units are linked by an ether linkage between the C2 and C4 of the ‘upper’ unit and the oxygen at the C7 position and the C6/C8 positions (respectively) of the lower unit, a Type A procyanidin is formed. Type A procyanidins are seen rarely when compared to Type B and C polyphenols.
Immunological Response to an Antigen
The immune system is a collection of mechanisms within a host that protects it against diseases by identifying and eliminating the pathogen. The system's response to a pathogen starts from the identification of a foreign protein to finally destroying the source of this foreign protein thereby protecting the host. Even the recognition of a simple protein from unicellular organism involves a series of complex steps which lead to the final elimination of the organism from the host. This entire process is the immunological response to the presence of a foreign protein or the antigen.
Resolution of infection by the immune system is the immunological response to the antigen, and it can be divided into 3 stages:
Activation and Mobilization: White Blood Cells (WBCs) are activated when they identify a foreign molecule or an antigen. Immune cells like macrophages and T cells release substances that attract other immune cells to the site of foreign molecule identified and thus mobilize the myriad of immune cells to eradicate the pathogen.
Regulation: The elicited immune response must be controlled in order to prevent excessive damage to the host. Regulator T-lymphocytes facilitate the control of the immune responses by secreting cytokines which act as the messengers of the immune system and thus regulate an exaggerated immune response.
Resolution: Infection resolution involves confining the pathogen and eliminating it from the body. After the pathogen is eliminated, most of the WBCs are destroyed, those that remain are called ‘memory cells’ and protect the host against future infection by the same pathogen by eliciting an early immune response to the pathogen.
A pathogen succeeds in causing an infection when the host is unable to surmount a defense strong enough to eliminate the pathogen. In such cases the antibodies produced by the host are insufficient to neutralize the existing numbers of the antigen. Hence the free antigens succeed in infecting the host. In such cases, external aids like antibiotics and antivirals are used to reduce the numbers of the antigen. Once the antigen numbers are reduced, the immunologic response is sufficient to eliminate the pathogen.
HIV Infection and AIDS:
Human immunodeficiency virus (HIV) is a retrovirus that destroys the immune system. This infection can eventually lead to Acquired Immunodeficiency Syndrome (AIDS), a serious and life-threatening condition in which the immune system fails to work properly. HIV primarily infects specific cells in the human immune system: “helper” T-lymphocytes (specifically CD4+ T cells), macrophages and dendritic cells. When CD4+ T cell numbers decline below a critical level, cell-mediated immunity is lost, and the body becomes progressively more susceptible to opportunistic infections.
HIV Life Cycle: Once HIV has entered the host, HIV needs specific host cells to facilitate its replication and propagation. The host cell in the case of HIV is the T-cell or CD4 cell.
1. Recognition of host and binding: HIV seeks out CD4 cells and attaches to them by way of a “lock and key” system via coreceptors on the cell surface. Proteins on the surface of HIV attach to complimentary proteins on the CD4 cell.
2. Attachment and entry into host: After attachment, the HIV injects viral proteins into the cellular fluids (cytoplasm) of the T-cell. This causes a fusion of the cell membrane to the outer envelope of the HIV.
3. Disassembly of viral proteins: In order to use its genetic material (RNA) for reproduction, the protective coating surrounding the RNA must be dissolved. Without this step, conversion of RNA to DNA (the building blocks of new HIV copies) cannot take place, and replication is halted.
4. Reverse transcription: Once inside the cell, the single stranded RNA of the HIV must be converted to the double stranded DNA. This step is brought about by the enzyme reverse transcriptase. Reverse transcriptase uses building blocks from the T-cell to help convert the viral RNA to DNA. The DNA contains the genetic information needed for HIV replication.
5. Replication and assembly into new virion: In order to replicate, the newly formed viral DNA must integrate into the host nucleus. This process is not entirely understood yet, but is believed to be aided by viral transport proteins. On integration, the virus gestates while the host cell prepares the proteins it requires to complete replication. Once the materials are available, they are cleaved by the virus based on requirement and structure and are subsequently assembled into new HIV. This process is aided by the protease enzyme.
6. Budding off from host cell: The final step of the viral replication cycle is called budding. With its genetic material tucked away and a new outer coat made from the host CD4 cell's membrane, the newly formed HIV pinches off and enters into circulation, ready to start the whole process again.
Current Interventions:
The current methods of interrupting HIV replication and propagation include: virus entry inhibitors; membrane fusion Inhibitors; and reverse transcriptase inhibitors; integrase inhibitors; protease inhibitors; maturation inhibitors, etc. The FDA has approved a number of drugs for treating HIV infection. Most of these drugs work by their antiretroviral (ARV) mechanism of action.
Infection with the Human Immunodeficiency Virus (HIV) presents political, economic, public health, social and scientific challenges to nations worldwide. At the close of 2007, an estimated 33.2 million people were living with HIV/AIDS worldwide. Hence there is an urgent need for the management and/or treatment of this disease with safer and more efficacious drugs. An additional challenge presented by this virus is its susceptibility to mutation. The viral proteins of HIV are prone to mutation and hence drug resistant strains pose an additional threat which creates a need for newer classes of drugs.
Influenza Virus:
Influenza is an infectious disease caused by RNA viruses of the family Orthomyxoviridae (the influenza viruses), that affects birds and mammals. Infection by this virus affects mainly the nose, throat, bronchi and, occasionally, lungs.
Structure of the Influenza Virus: The Influenza virus is classified into 3 categories: Influenza virus A, B and C. The 3 subtypes of influenza viruses have a very similar overall structure. The viruses are made of a viral envelope containing two main types of glycoproteins which are wrapped around a central core. The central core contains the viral RNA genome and other viral proteins that package and protect this RNA. Hemagglutinin (HA) and neuraminidase (NA) are the two large glycoproteins on the outside of the viral particles.
Influenza Virus A: The type A viruses are the most virulent human pathogens among the three influenza types and cause the most severe disease. The influenza A virus can be subdivided into different serotypes based on the antibody response to these viruses. The serotypes that have been confirmed in humans, ordered by the number of known human pandemic deaths, are: H1N1, H2N2, H3N2, H5N1, H7N7, H1N2, H9N2, H7N2, H7N3, H10N7. Influenza A viruses have caused several pandemics during the last century, and continue to cause annual epidemics. The emergence of new strains of influenza continues to pose challenges to public health and scientific communities. The H1N1 virus is a serotype of the Influenza A virus and is one of the most virulent strains known to affect human beings. The H1N1 serotype has been responsible for millions of deaths in 1918 (Spanish Flu) and is more recently causing a Swine Flu global pandemic.
Influenza Viral Life Cycle: The influenza viral replication and propagation process is outlined below:
1. Recognition of host and binding: Binding of the virus to the host cell using the HA protein to sialic acid bound to sugars on the surfaces of epithelial cells. Epithelial cells are typically present in the nose, throat and lungs of mammals and in the intestines of birds
2. Attachment and entry into host: After the binding, the HA protein is cleaved off and the virus enters the cell by endocytosis.
3. Disassembly of viral proteins: Once the virus enters the cell, the pH and ambient conditions of the endosome lead to:                a. A part of HA fusing the viral envelope to the vacuole membrane        b. The M2 ion channel permits the entry of protons to the viral core which acidify the viral core leading to its disassembly and the subsequent release of the viral RNA and core proteins into the host cell cytoplasm        
4. Reverse transcription: The viral RNA and core proteins are now transported into the cell nucleus where the RNA is transcribed and further translated into viral proteins.
5. Budding off from host cell: HA and NA proteins form clusters near the cell membrane which subsequently also house the viral RNA and core proteins, which then lead to ‘budding’ of the virus and propagation for subsequent infection.
As seen from the infection and propagation steps detailed above HA and NA play an important role in infection. Before release of the virion, NA also cleaves sialic acid so as to prevent the binding of HA to sialic acid.
Current Interventions for Influenza A virus: There are two classes of drugs approved by the United States FDA against the Influenza A virus: Ion channel inhibitors like Adamantanes (amantadine hydrochloride and rimantadine); and Neuraminidase inhibitors like Oseltamivir (TAMIFLU) and Zanamivir (RELENZA)
The Influenza A virus is prone to mutations. These mutations are primarily of viral proteins like NA, HA and M2 ion channel proteins, and hence inhibitors of these proteins will be ineffective against mutant strains. The mutation potential and the 2009 Influenza A global pandemic presents an urgent need for therapies which offer a treatment and prevention options against this virus.
PRIOR ART
Richard Anderson et al, “Isolation and characterization of polyphenols Type A polymers from cinnamon with insulin-like biological activity” in the Journal of Agricultural and Food Chemistry, 2004, pp 52, 65-70.
This paper describes an aqueous extract of commercial cinnamon and has identified polyphenolic polymers that increase glucose metabolism by roughly 20 fold in in vitro cell lines. They have used Cinnamomum cassia (Korintji cassia) for the preparation of this extract. This variety has a high content of coumarin and cinnamaldehyde.
This paper further describes a preparatory HPLC method for the preparation and characterization of this aqueous extract.
This publication describes A-type doubly linked procyanidin of catechins. This paper has identified trimer (molecular weight 864), tetramer (molecular weight 1152) and oligomer of catechins that are isolated from cinnamon.
Kilkuskie et al, “HIV and reverse transcriptase inhibition by tannins” in the Bioorqanic and Medicinal Chemistry Letters, 1992, Vol 2, pp 1529-1534.
This publication evaluates tannins and condensed tannins for their anti-HIV activity and their potential to inhibit the reverse transcriptase enzyme. Although this study discovered some tannins with anti-HIV activity, they were burdened with the associated toxicities. This publication talks about 3 compounds which are condensed forms of catechins. Molecules 40, 44 and 45 are dimers, trimers and tetramers of catechins. This paper concluded that there was no correlation between the inhibition of the RT enzyme and the anti-HIV action of these tannins. Additionally, Molecules 44 and 45 showed an anti-HIV activity of 90% and 73% respectively, but did not show significant inhibition of the RT enzyme.
Michael Ovadia et al. in Patent Application US 2006 275515A1
This patent titled “Anti-viral preparations obtained from a natural cinnamon extract” have described a natural aqueous extract obtained from cinnamon having anti-viral properties. This document describes a water extract of cinnamon which is subjected to salting out precipitation with a salt. This precipitate is redissolved in water or buffer and purified using sepharose chromatography and subsequently eluted with another buffer and galactose.
The commonly used process of salting out refers to the selection of high molecular weight molecules (usually peptides). Hence it is quite evident from this process that the process described in this document is aimed at recovering high molecular weight molecules (about 10 Kda).
The active ingredient of the composition as per the claim is having a molecular weight more than 10 KDA and it responds to an absorbance at 280 nm at between about 15 and 20OD. This compound is finally eluted from the sepharon column using phosphate buffer and galactose. Therefore, the final compound will have high concentrations of phosphates and galactose.
This high molecular weight compound described in this application has been tested in influenza A PR 8 virus, Para Influenza (Sendai) virus, Pre-absorption into erythrocytes and weight gain in mice infected with influenza or the sendai virus and an HIV syncytia study.
Example 13 of this patent application describes a test done with this extract in MT2 cells to check the effect on Syncytia formation. As per FIG. 15 of this application at concentrations of 60 to 100 micrograms, it inhibits syncytia formation. Syncytia formation is not a confirmatory test for anti-viral activity. This is elucidated with evidence in the following publication. [Gueseppe pantaleo et al Eur J immunology 1991, 21, 1771:1774 dissociation between syncytia formation and HIV spreading. Suppressing Syncytia formation does not necessarily reflect inhibition of HIV infection.]
Although, the disclosed extract showed potential to inhibit syncytia formation, it should be noted that only some strains of HIV cause syncytia formation. Additionally syncytia formation cannot be linked to the presence or to the progression of HIV infection or AIDS. Syncytia formation is merely a phenotype that may be expressed by some strains. Lack of syncytia formation cannot be linked to the absence of HIV or to the management of the infection.